Passive acoustic sensors have been used to analyze characteristic acoustic energy emissions generated by inorganic chemical reactions and chemical changes. Some of these chemical changes which produce acoustic energy include liquid-solid and solid-liquid transitions, dissolution, hydration and gelation. Chemical changes are normally accompanied by energy transfer and some of the energy released may be in the form of acoustic waves. Betteridge et al. examined acoustic energy spectra from 43 different chemical systems. They found a correlation between chemical and acoustic events, although some acoustic responses were difficult to explain. They speculated that the thermal changes occurring during chemical reactions were sources of acoustics signals. D. Betteridge et al., "Acoustic Emissions From Chemical Reactions", Anal. Chem. Vol 53, p. 1064-1073 (1981). Sawada and Abe investigated acoustic emissions arising from a gelation of sodium carbonate and calcium chloride. They obtained several acoustic signals during the gelation reaction. They explained the initial acoustic energy signals as thermally caused via exothermic reaction in the gel formation, but interpretation of the other responses needed further experimental work. Sawada and Abe, "Acoustic Emissions Arising From the Gelation of Sodium Carbonate and Calcium Chloride", Anal. Chem. Vol 5, pp 366-367 (1985). Acoustic emission from phase transitions of four chemical systems was investigated by Sawada et al. They found that acoustic energy signals were influenced by changes of volume, heat balance, and reaction rate. Sawada et al., "Acoustic Emission From Phase Transition of Some Chemicals", Anal. Chem. Vol 57, pp 1743-1745 (1985).
A model process, the hydration of silica gel, was investigated in detail by Belchamber et al. using acoustic energy emissions. They found that the hydration process for the silica was a two-step process and obtained important information about the kinetics of that process. Their results suggest that acoustic energy monitoring could be an effective way of gaining often unique information about difficult-to-study chemical processes and could be applied for monitoring chemical processes. Belchamber et al., Anal. Chem. Vol 58, p. 1877 (1986).
Wentzel and Wade showed that chemical reactions have their own unique acoustic energy signatures. They analyzed frequency spectra of the acoustic energy signal for several chemical systems using Fourier transform technique. They also found that the different physical processes yield different contributions to acoustic energy response, ie. gas release is shown to result in low-frequency signals, while crystal fracture produces higher frequency range acoustic energy. Wentzel and Wade, "Chemical Acoustic Emission Analysis in Frequency Domain", Anal. chem. vol 61, pp 2638-2642 (1989). Wentzel et al. found that different bubble evolution sites on an enzyme catalytic surface produced repeatable emission wave fronts, and each had its own particular acoustic signature. Wentzel et al. used the enzyme catalase immobilized on the transducer surface to accelerate the conversion of hydrogen peroxide to gas. Wentzel et al. "Evaluation of Acoustic Emission as a Means of Quantitative Chemical Analysis", Anal. chem. Acta, vol 246, pp 43-53 (1991).
Recently, Lec et al. investigated the kinetics of the causticization process using the acoustic energy technique. The causticization process consists of two reactions, a lime slaking reaction and a causticization reaction. It was found that the acoustic energy frequency spectra of these reactions were different. Also, a new mechanism, caused by the ion--ion exchange reaction during the causticization process, has been identified via specific acoustic energy signals. Lec et al., Proc. SPIE Smart Sensing, Processing, & Instrumentation, vol 1918, pp 440-448 (1993) and S. W. Bang, R. M. Lec, J. M. Genco et al., "Acoustic Emission Chemical Sensor", IEEE 1993 Ultrasonics symposium, (Nov. 2-5, 1993).
Generally, acoustic sensors can be categorized as either active or passive. Active sensors transmit an acoustic wave into a medium under test, the acoustic wave interacts with the medium, and measured parameters of the acoustic wave are correlated with medium properties. Passive acoustic sensors measure acoustic signals which are generated in the medium itself during physical or chemical processes.
Research on active acoustic sensors in the biochemical and biomedical areas has occurred for several decades. Active acoustic sensors have been used to detect organic gases, viscosity or ion concentration in solutions. In recent years active acoustic sensors have been used for monitoring immunological reactions. To date, only three types of acoustic devices, a quartz crystal resonator (QCR), the acoustic plate mode (APM) device and the bulk wave (SSBW) device have been used for immunoassay purposes. The methods all involved the surface immobilization of one of the biocomponents, usually the antibody, and use of active acoustic probes.
While active acoustic sensor technology has now been used in biochemical and biological reactions, it has not been conceived to use passive acoustic sensor technology to detect the subtleties of biological reactions such as the binding reactions of biorecognition molecules. It is established that the making and breaking of chemical bonds during chemical reactions generates sufficient acoustic energy emission for distinguishing the chemical changes by the characteristic frequency spectrum of the acoustic emissions over time. It is not obvious or suggested however, that the binding reactions of biorecognition molecules such as antigen/antibody reactions and complementary nucleic acid reactions cause sufficient acoustic emissions and in distinguishing characteristic patterns for discriminating between such similar binding reactions.
Immunoassays are well-known for their selectivity and speed because of the inherent specificity and high binding constants of antibodies. The most desirable immunoassay is one which is homogeneous. In such an assay, the patient specimen containing an unknown level of analyte is mixed directly with the antibody reagent. After some incubation period, a direct result is obtained in the form of a visual observation or instrument measurement of fluorescence, color, chemiluminescence etc. Commercial homogeneous immunoassays can thus be used or added to existing clinical analyzers and are much in demand due to the ease of use. Examples of such assay technologies include Syva's EMIT (TM) assays that are enzyme mediated and Abbott's TDX (TM) that are fluorescence polarization based. These systems are designed to quantitate analytes such as drugs and small molecular weight hormones.
However, with the exception of nephelometry, there are no other homogeneous immunoassay systems which are capable of quantitating detected macromolecules such as proteins. Those immunoassays which do, are heterogenous, requiring multiple separation and wash steps, followed ultimately by a "reporter" system such as an isotope (RIA) or enzyme (ELISA). These later assays, though sensitive, require a minimum assay time of at least 2-3 hours and are discrete, i.e. with single, end-point determination.
Immunonephelometry is used commercially to quantitate clinically significant human proteins and exhibits the inherent simplicity and ease of use of a true homogeneous assay. Specific antiserum is mixed with a dilution of patient serum and the resulting turbidity which develops over time is measured optically by absorbance (turbidimetry) or reflective fluorescence (nephelometry). However, this technology has several major drawbacks. First, the specimen to be tested must have optical clarity i.e. not be turbid or contain solids. Second, the reaction cell or cuvette requires a high quality optical window and usually two. Third, the lower limit of sensitivity is .sup.18 10 .mu.g/ml of antigen protein.
These drawbacks have meant that this technique has been limited in its application to the measurement of selected serum/plasma proteins found in the higher concentrations as specified above. Thus there are a number of clinically significant protein analytes such as tumor markers, ferritin, and .beta.2-microglobulin which are present at lower concentrations and therefore must be assayed by other more tedious and time consuming techniques such as the enzyme linked immunosorbent assay i.e. ELISA. Because of the requirement for specimen clarity, immunoturbidimetry cannot be applied to industrial use. Thus application areas such as food processing are not possible with current homogeneous assays.